Gamma-aminobutyric acid (GABA) is the major inhibitory neurotransmitter in the nervous system. Alterations in GABA neurotransmission have been implicated in the etiology of several human neurological and psychiatric disorders. The synthesis of GABA is catalyzed by the enzyme glutamate decarboxylase (GAD) in vertebrates and invertebrates. Using a cloned mammalian GAD cDNA as a probe, we have isolated homologous cDNA and genomic DNA clones from Drosophila melanogaster. The clones appear to represent a single gene which is located in polytene chromosome region 64A. Limited cDNA sequencing demonstrates that the Drosophila gene encodes a GAD-homologous protein. In order to initiate developmental and molecular studies of GABA neurotransmitter metabolism, the following investigations are proposed. (1) The complete sequence of the Drosophila GAD- homologous protein will be deduced from sequence of cDNA clones. Gene dosage and protein expression techniques will be employed to verify that the Drosophila gene encodes functional GAD protein. (2) Distinct forms of GAD have been documented in mammals and insects, including Drosophila. As a prelude to studying the expression and functional significance of GAD isoforms, we will determine how many genes encode the distinct forms of enzyme. 3) The normal ontogeny and anatomical distribution of GABAergic neurons will be determined by examining the developmental and tissue expression of GAD gene RNA as well as GAD and GABA immunoreactivity. (4) Unconditional and temperature-sensitive GAD mutations will be isolated for use in genetic studies of GABAergic systems. (5) Temperature-sensitive GAD mutations will be used to eliminate GAD activity at specified developmental stages in order to assess the effects of GABA deficits on the differentiation and/or maintenance of GABAergic and other neural pathways.